The present invention relates to oxygenated esters of 4-substituted-phenylamino benzhydroxamic acid derivatives, pharmaceutical compositions, and methods of use thereof.
MAPK/ERK Kinase (xe2x80x9cMEKxe2x80x9d) enzymes are dual specificity kinases involved in, for example, immunomodulation, inflammation, and proliferative diseases such as cancer and restenosis.
Proliferative diseases are caused by a defect in the intracellular signaling system, or the signal transduction mechanism of certain proteins. Defects include a change either in the intrinsic activity or in the cellular concentration of one or more signaling proteins in the signaling cascade. The cell may produce a growth factor that binds to its own receptors, resulting in an autocrine loop, which continually stimulates proliferation. Mutations or overexpression of intracellular signaling proteins can lead to spurious mitogenic signals within the cell. Some of the most common mutations occur in genes encoding the protein known as Ras, a G-protein that is activated when bound to GTP, and inactivated when bound to GDP. The above-mentioned growth factor receptors, and many other mitogenic receptors, when activated, lead to Ras being converted from the GDP-bound state to the GTP-bound state. This signal is an absolute prerequisite for proliferation in most cell types. Defects in this signaling system, especially in the deactivation of the Ras-GTP complex, are common in cancers, and lead to the signaling cascade below Ras being chronically activated.
Activated Ras leads in turn to the activation of a cascade of serine/threonine kinases. One of the groups of kinases known to require an active Ras-GTP for its own activation is the Raf family. These in turn activate MEK (e.g., MEK1 and MEK2) which then activates the MAP kinase, ERK (ERK1 and ERK2). Activation of MAP kinase by mitogens appears to be essential for proliferation; constitutive activation of this kinase is sufficient to induce cellular transformation. Blockade of downstream Ras signaling, for example, by use of a dominant negative Raf-1 protein, can completely inhibit mitogenesis, whether induced from cell surface receptors or from oncogenic Ras mutants. Although Ras is not itself a protein kinase, it participates in the activation of Raf and other kinases, most likely through a phosphorylation mechanism. Once activated, Raf and other kinases phosphorylate MEK on two closely adjacent serine residues, S218 and S222 in the case of MEK-1, which are the prerequisite for activation of MEK as a kinase. MEK in turn phosphorylates MAP kinase on both a tyrosine, Y185, and a threonine residue, T183, separated by a single amino acid. This double phosphorylation activates MAP kinase at least 100-fold. Activated MAP kinase can then catalyze the phosphorylation of a large number of proteins, including several transcription factors and other kinases. Many of these MAP kinase phosphorylations are mitogenically activating for the target protein, such as a kinase, a transcription factor, or another cellular protein. In addition to Raf-1 and MEKK, other kinases activate MEK, and MEK itself appears to be a signal integrating kinase. Current understanding is that MEK is highly specific for the phosphorylation of MAP kinase. In fact, no substrate for MEK other than the MAP kinase, ERK, has been demonstrated to date and MEK does not phosphorylate peptides based on the MAP kinase phosphorylation sequence, or even phosphorylate denatured MAP kinase. MEK also appears to associate strongly with MAP kinase prior to phosphorylating it, suggesting that phosphorylation of MAP kinase by MEK may require a prior strong interaction between the two proteins. Both this requirement and the unusual specificity of MEK are suggestive that it may have enough difference in its mechanism of action to other protein kinases that selective inhibitors of MEK, possibly operating through allosteric mechanisms rather than through the usual blockade of the ATP binding site, may be found.
It has been found that the compounds of the present invention are inhibitors of MEK and are useful in the treatment of a variety of proliferative disease states, such as conditions related to the hyperactivity of MEK, as well as diseases modulated by the MEK cascade.
The present invention provides a compound of Formula 
wherein
W is 
R2 is hydrogen, methyl, fluorine, or chlorine;
R3 is hydrogen or fluorine;
R4 is xe2x80x94CH2S(CH2)m(CH3), xe2x80x94Cxe2x89xa1Cxe2x80x94(CH2)qNHCH3, xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3, (Z)xe2x80x94CHCHCH2OCH3, xe2x80x94(CH2)nCO2R6, xe2x80x94(CF2)pCF3, xe2x80x94CH2(CF2)qCF3, xe2x80x94(CH2)mCF(CF3)2, xe2x80x94CH(CF3)2, xe2x80x94CF2CF(CF3)2, xe2x80x94C(CF3)3, xe2x80x94Cxe2x89xa1Cxe2x80x94(CH2)qN(CH3)2, xe2x80x94(Z)xe2x80x94CHCHxe2x80x94(CH2)qNHCH3, (Z)xe2x80x94CHCHxe2x80x94(CH2)qN(CH3)2, or C(O)C1-3 alkyl;
m is 0 to 1;
n is 0 to 2;
p is 1 to 5;
q is 1 to 2;
R5 is hydrogen, fluorine, bromine, or chlorine;
R6 is hydrogen, methyl or ethyl;
and pharmaceutically acceptable salts, (C1-6) amides and (C1-6) esters thereof.
The present invention provides compounds of Formula I wherein W is 
R2 is hydrogen, fluorine, or chlorine; R3 is fluorine; R4 is xe2x80x94(CH2)nCO2R6 where n is 0; R4 is xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3; or R4 is C(O)C1-3 alkyl; and R5 is hydrogen.
The invention also provides a pharmaceutical composition comprising a compound of Formula I and a pharmaceutically acceptable carrier.
Additionally, the invention provides a method of treating a proliferative disease in a patient in need thereof comprising administering a therapeutically effective amount of a compound of Formula I.
The invention also provides the use of a compound of Formula I for the manufacture of a medicament for the treatment of a proliferative disease.
Furthermore, the invention provides methods of treating cancer, restenosis, psoriasis, autoimmune disease, atherosclerosis, osteoarthritis, rheumatoid arthritis, heart failure, chronic pain, and neuropathic pain in a patient in need thereof comprising administering a therapeutically effective amount of a compound of Formula I.
The invention also provides the use of a compound of Formula I for the manufacture of a medicament for the treatment of cancer, restenosis, psoriasis, autoimmune disease, atherosclerosis, osteoarthritis, rheumatoid arthritis, heart failure, chronic pain, and neuropathic pain.
In addition, the invention provides a method for treating cancer in a patient in need thereof comprising administering a therapeutically effective amount of a compound of Formula I in combination with radiation therapy or at least one chemotherapeutic agent.
Certain terms are defined below and by their usage throughout this disclosure.
The terms xe2x80x9chalogenxe2x80x9d or xe2x80x9chaloxe2x80x9d in the present invention refer to a fluorine, bromine, chlorine, and iodine atom or fluoro, bromo, chloro, and iodo. The terms fluorine and fluoro, for example, are understood to be equivalent herein.
Alkyl groups, such as xe2x80x9cC1-6 alkylxe2x80x9d, include aliphatic chains (i.e., hydrocarbyl or hydrocarbon radical structures containing hydrogen and carbon atoms) with a free valence. Alkyl groups are understood to include straight chain and branched structures. Examples include methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, t-butyl, pentyl, 2-pentyl, 3-pentyl, isopentyl, neopentyl, (R)-2-methylbutyl, (S)-2-methylbutyl, 3-methylbutyl, 2,3-dimethylpropyl, hexyl, and the like. The term xe2x80x9cC1-6 alkylxe2x80x9d includes within its definition the terms xe2x80x9cC1-4 alkylxe2x80x9d and xe2x80x9cC1-2 alkylxe2x80x9d.
Alkenyl groups are analogous to alkyl groups, but have at least one double bond (two adjacent sp2 carbon atoms). Depending on the placement of a double bond and substituents, if any, the geometry of the double bond may be entgegen (E), or zusammen (Z), cis, or trans. Similarly, alkynyl groups have at least one triple bond (two adjacent sp carbon atoms). Unsaturated alkenyl or alkynyl groups may have one or more double or triple bonds, respectively, or a mixture thereof. Like alkyl groups, unsaturated groups may be straight chain or branched. Examples of alkenyls and alkynyls include vinyl, allyl, 2-methyl-2-propenyl, cis-2-butenyl, trans-2-butenyl, and acetyl.
The present invention includes the hydrates and the pharmaceutically acceptable salts and solvates of the compounds defined by Formula I. The compounds of this invention can possess a sufficiently basic functional group, and accordingly react with any of a number of inorganic and organic acids, to form a pharmaceutically acceptable salt.
The term xe2x80x9cpharmaceutically acceptable saltxe2x80x9d as used herein, refers to salts of the compounds of Formula I which are substantially nontoxic to living organisms. Typical pharmaceutically acceptable salts include those salts prepared by reaction of the compounds of the present invention with a pharmaceutically acceptable mineral or organic acid. Such salts are also known as acid addition salts. Such salts include the pharmaceutically acceptable salts listed in J of Pharm Sci. 1977;66:2-19, which are known to the skilled artisan.
Acids commonly employed to form acid addition salts are inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and the like, and organic acids such as p-toluenesulfonic, methanesulfonic acid, benzenesulfonic acid, oxalic acid, p-bromophenylsulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like. Example of such pharmaceutically acceptable salts are the sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, bromide, hydrobromide, iodide, acetate, propionate, decanoate, caprate, caprylate, acrylate, ascorbate, formate, hydrochloride, dihydrochloride, isobutyrate, caproate, heptanoate, propiolate, glucuronate, glutamate, propionate, phenylpropionate, salicylate, oxalate, malonate, succinate, suberate, sebacate, fumarate, malate, maleate, hydroxymateate, mandelate, mesylate, nicotinate, isonicotinate, cinnamate, hippurate, nitrate, stearate, phthalate, teraphthalate, butyne-1,4-dioate, butyne-1,4-dicarboxylate, hexyne-1,4-dicarboxylate, hexyne-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, hydrozybenzoate, methoxybenzoate, dinitrobenzoate, o-acetoxybenzoate, naphthalene-2-benzoate, phthalate, p-toluenesulfonate, p-bromobenzenesulfonate, p-chlorobenzenesulfonate, xylenesulfonate, phenylacetate, trifluoroacetate, phenylpropionate, phenylbutyrate, citrate, lactate, xcex1-hydroxybutyrate, glycolate, tartrate, hemitartrate, benzenesulfonate, methanesulfonate, ethanesulfonate, propanesulfonate, hydroxyethanesulfonate, 1-naphthalenesulfonate, 2-naphthalenesulfonate, 1,5-naphthalenedisulfonate, mandelate, tartarate, and the like. A preferred pharmaceutically acceptable salt is hydrochloride.
It should be recognized that the particular counterion forming a part of any salt of this inventions is usually not of a critical nature, so long as the salt as a whole is pharmacologically acceptable and as long as the counterion does not contribute undesired qualities to the salt as a whole. It is further understood that such salts may exist as a hydrate.
As used herein, the term xe2x80x9cstereoisomerxe2x80x9d refers to a compound made up of the same atoms bonded by the same bonds but having different three-dimensional structures which are not interchangeable. The three-dimensional structures are called configurations. As used herein, the term xe2x80x9cenantiomerxe2x80x9d refers to each of two stereoisomers whose molecules are nonsuperimposable mirror images of one another. The term xe2x80x9cchiral centerxe2x80x9d refers to a carbon atom to which four different groups are attached. As used herein, the term xe2x80x9cdiastereomersxe2x80x9d refers to stereoisomers which are not enantiomers. The terms xe2x80x9cracematexe2x80x9d or xe2x80x9cracemic mixturexe2x80x9d refer to a mixture of enantiomers.
The enantiomers of compounds of the present invention can be resolved by one of ordinary skill in the art using standard techniques well-known in the art, such as those described by J. Jacques, et al., xe2x80x9cEnantiomers, Racemates, and Resolutionsxe2x80x9d, John Wiley and Sons, Inc. 1981. Examples of resolutions include recrystallization techniques or chiral chromatography.
Some of the compounds of the present invention have one or more chiral centers and may exist in a variety of stereoisomeric configurations. As a consequence of these chiral centers, the compounds of the present invention occur as racemates, mixtures of enantiomers and as individual enantiomers, as well as diastereomers and mixtures of diastereomers. All such racemates, enantiomers, and diastereomers are within the scope of the present invention.
The compounds of Formula I can be prepared by techniques and procedures readily available to one of ordinary skill in the art, for example, by following the procedures as set forth in the following Schemes, or analogous variants thereof. These synthetic strategies are further exemplified in Examples below. These Schemes are not intended to limit the scope of the invention in any way.
As used herein, the following terms have the meanings indicated: xe2x80x9cBOCxe2x80x9d refers to tert-butoxycarbonyl; Celite(copyright) refers to a filter agent which is acid washed and approximately 95% SiO2; xe2x80x9cDMAxe2x80x9d refers to N,N-dimethylacetamide; xe2x80x9cDMT-MMxe2x80x9d refers to 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride; xe2x80x9cEtOAcxe2x80x9d refers to ethyl acetate; xe2x80x9cEtOHxe2x80x9d refers to ethanol; xe2x80x9cEt2Oxe2x80x9d refers to diethyl ether; xe2x80x9chxe2x80x9d refers to hours; xe2x80x9cLiHMDSxe2x80x9d refers to lithium 1,1,1,3,3,3-hexamethyldisilazane or lithium bis(trimethylsilylamide); xe2x80x9cLindlar catalystxe2x80x9d refers to a Pd/CaCO3 catalyst washed with Pb(OAc)2; xe2x80x9cMexe2x80x9d refers to methyl; xe2x80x9cMeOHxe2x80x9d refers to methanol; xe2x80x9cMsClxe2x80x9d refers to methane sulfonyl chloride; xe2x80x9cPd/Cxe2x80x9d refers to palladium on carbon; xe2x80x9c(Ph3P)2PdCl2xe2x80x9d refers to dichlorobis(triphenylphosphine)palladium(II); xe2x80x9c(Ph3P)4Pdxe2x80x9d refers to tetrakis(triphenylphosphine)palladium(0); xe2x80x9cPyBopxe2x80x9d refers to benzotriazole-1-yl-oxy-tris-pyrrolidino-phosphonium hexafluorophosphate; xe2x80x9cRTxe2x80x9d refers to room temperature; xe2x80x9cTEAxe2x80x9d refers to triethylamine; xe2x80x9cTFAxe2x80x9d refers to trifluoroacetic acid; xe2x80x9cTHFxe2x80x9d refers to xe2x80x9ctetrahydrofuran; xe2x80x9cTLCxe2x80x9d refers to thin layer chromatography; and xe2x80x9cTMSxe2x80x9d refers to trimethylsilyl. All other terms and substituents, unless otherwise indicated, are previously defined. The reagents and starting materials are readily available to one of ordinary skill in the art. Schemes 1 and 2 provide syntheses of the compounds of Formula I. 
In Scheme 1, Step A, a 2-(arylamino)-benzoic acid or diphenylamine (3) is prepared from the coupling of a suitable benzoic acid (1) and a suitable aniline (2) in the presence of a strong base, for example, lithium 1,1,1,3,3,3-hexamethyldisilazane (LiHMDS), in a polar aprotic solvent such as tetrahydrofuran, acetonitrile or dimethylformamide. For example, the aniline (2) and the benzoic acid (1) are dissolved in a suitable organic solvent and cooled to about xe2x88x9278xc2x0 C. under nitrogen. The suspension is treated with an excess of a suitable base, such as LiHMDS, and allowed to warm to room temperature. The reaction is typically complete within about 2 hours to about 5 days. The resulting benzoic acid (3) can be isolated by removing the solvent, for example by evaporation under reduced pressure or by filtering the precipitated solid through Celite(copyright) and washing with a suitable solvent. The benzoic acid (3) can be further purified, if desired, by standard methods such as chromatography, crystallization, or distillation.
In Scheme 1, Step B, the compounds of Formula I are generally obtained by the union of 2-(arylamino)-benzoic acid (3) with an alkoxylamine (4) by the action of a peptide coupling agent in the presence of a base, if necessary. It is understood that the alkoxylamine (4) may be suitably protected. In such instances, Scheme 1 may be modified to include a removal of the protecting group by a procedure known in the art. Preferred coupling agents include 1,1xe2x80x2-carbonyldiimidazole (CDI), lithium bis (trimethylsilylamide) (LiHMDS), diphenylphosphinic chloride (DPP-Cl), benzotriazol-yl-oxy-tripyrolidinophosphonium hexafluorophosphate (PyBOP), benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate (BOP), N,Nxe2x80x2-dicyclohexylcarbodiimide (DCC), or 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDCI). Preferred bases include diisopropylethylamine, triethylamine, 4-methylmorpholine, or pyridine or a substituted pyridine, for example, 4-dimethyaminopyridine or 2,6-dimethylpyridine. Preferred solvents are polar aprotic solvents such as dichloromethane, tetrahydrofuran, or dimethylformamide. The reactions are generally carried out at a temperature between about xe2x88x9278xc2x0 C. to about 25xc2x0 C., and are normally complete within about 1 hour to about 5 days. The product amide can be isolated by removing the solvent, for example by evaporation under reduced pressure, and further purified, if desired, by standard methods such as chromatography, crystallization, or distillation.
It would be understood by one of skill in the art that the substituent at R4 on the diphenylamine (3) or the substituent at R4 on the compound of Formula I can be reduced before the coupling reaction. The reduction is performed on alkene or alkyne derivatives under conditions known in the art, such as through hydrogenation, for example with Pd/C under an atmosphere of hydrogen.
Alternately, the compounds of formula I are generally prepared as shown in Scheme 1, steps C and D by the contact of alkoxyamine (4) with xe2x80x9cactivatedxe2x80x9d benzoic acid derivatives (3a), wherein the activating group xe2x80x9cXxe2x80x9d completes an acid halide, anhydride, mixed anhydride, or an activated ester, such as a pentafluorophenyl ester, nitrophenyl ester or thioester. Preferred bases include diisopropylethylamine, triethylamine, 4-methylmorpholine, imidazole, pyridine or a substituted pyridine, for example, 4-dimethyaminopyridine or 2,6-dimethylpyridine. Preferred solvents are polar aprotic solvents such as dichloromethane, tetrahydrofuran, or dimethylformamide. 
In Scheme 2, Step A, a 4-iodo phenylamino benzoic acid (3xe2x80x2) is prepared from the union of a suitable benzoic acid (1) and a suitable 4-iodoaniline (5), according to the general procedure of Scheme 1, Step A.
In Scheme 2, Step B, the 4-iodo phenylamino benzoic acid (3xe2x80x2) is coupled with an alkoxylamine (4) according to the general procedure of Scheme 1, Step B or Scheme 1, Steps C and D.
In Scheme 2, Step C, the compounds of Formula I are prepared from the 4-iodo-phenylamino benzamide (6), by transition metal-promoted coupling with reagent M-R4 (7) in a suitable solvent such as triethylamine. The entire mixture is stirred from about 2 to 24 hours at room temperature. The transition metal-promoted coupling may be carried out with a palladium(0) or palladium (II) coupling agent, such as (Ph3P)4Pd or (Ph3P)2PdCl2 and cuprous iodide. M is defined as a functional group known to transfer a carbon radical fragment in transition metal-promoted coupling processes. Examples of a suitable M group include trialkylstannyl, trialkylsilyl, trimethylsilyl, zinc, copper, boron, magnesium and lithium. It would be understood by one of skill in the art that the substituent R4 may be further transformed, such as by oxidation, reduction, deprotection, or hydrogenation. The substituent R4 may also be transformed into a different R4 through standard synthetic procedures known to one of skill in the art. The resulting compound of formula I, as well as the protected Formula I compound, can be isolated by removing the solvent, for example by evaporation under reduced pressure, and further purified, if desired, by standard methods such as chromatography, crystallization, or distillation.
The aniline (2) can be prepared by techniques and procedures readily available to one of ordinary skill in the art and by following the procedures as set forth in the following Schemes, or analogous variants thereof. These Schemes are not intended to limit the scope of the invention in any way. 
In Scheme 3, Step A, a suitable thiolate (11) is reacted with a 4-tert-butoxycarbonylamino-3-substituted-benzyl bromide, such as 4-tert-butoxycarbonylamino-3-fluorobenzyl bromide (J Med Chem. 2000;43:5017) to provide a compound of structure (12). In Step B, the BOC protecting group of compound (12) is hydrolyzed with, for example, TFA to provide the desired aniline (2a). 
In Scheme 4, Step A, a suitable acetylene (14) such as methylpropargyl ether, and a suitable 4-iodoaniline (13) are coupled via the Sonogashira reaction. For example, a 4-iododaniline (13), such as 2-fluoro-4-iodoaniline, is combined with CuI and (Ph3P)2PdCl2 under nitrogen. An acetylene derivative (14) is added in a suitable solvent, such as TEA, and the entire mixture is stirred for about 2 to 24 hours at room temperature. 
After reaction of the aniline (2b) with a suitable benzoic acid (1) under the standard conditions to form the diphenylamine, as in Scheme 1, Step A above, the alkynyl diphenylamine (3a) is dissolved in a suitable solvent, such as tetrahydrofuran, in the presence of a catalyst, such as Lindlar catalyst or palladium on carbon and, if desired, a suitable compound which disrupts the action of the catalyst, such as quinoline or pyridine. The mixture is stirred under an atmosphere of hydrogen from about 1 to 24 hours at room temperature. The resulting alkenyl diphenylamine (3b) can be isolated by removing the solvent, for example by evaporation under reduced pressure, and further purified, if desired, by standard methods such as chromatography, crystallization, or distillation. Reaction with a suitable alkoxylamine, with the 2-(arylamino)benzoic acids (3a and 3b) under the conditions of Scheme 1, Step B gives the desired compounds of Formula I, wherein R4 is xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3 or xe2x80x94Cxe2x95x90Cxe2x80x94CH2OCH3. 
In Scheme 6, Step B, a suitable acetylene derivative (16), which has been protected, for example with a tetrahydropyranyl ether, and a suitable 4-iodoaniline (13) such as 2-fluoro-4-iodoaniline are coupled via the Sonogashira reaction as in Scheme 4, Step A, to provide the desired aniline (2c). 
In Scheme 7, Step A, the propargyl alcohol compound (20) is prepared from the iodide compound (18) by transition metal-promoted coupling according to the general procedure of Scheme 2, Step C.
In Scheme 7, Step B, the propargyl alcohol compound (20) is converted to a fully saturated substituent through hydrogenation, for example with Pd/C under an atmosphere of hydrogen to provide the methyl 4-substituted-phenylamino benzoate (22).
In Scheme 7, Step C, methyl 4-substituted-phenylamino benzoate (22) is deprotected in a manner known to one of skill in the art, for example, aqueous NaOH in EtOH, then coupled with an hydroxylamine according to the general procedure of Scheme 1, Step B.
In Scheme 7, step D, the compound (24) is dissolved in a suitable solvent such as tetrahydrofuran and reacted with methanesulfonyl chloride to give the intermediate mesylate, then NaI in EtOAc to give the iodide compound (26).
In scheme 7, steps E and F, the iodide compound (26) is reacted with methylamine and dimethylamine respectively to give compounds of formula I wherein m is 3 and R4 is xe2x80x94(CH2)mNHCH3 (28) and xe2x80x94(CH2)mN(CH3)2 (30). 
In Scheme 8, Step A, the desired perfluoroalkyl anilines (2d) are prepared by an Ullmann condensation of a suitable 4-iodoaniline (5), such as 2-fluoro-4-iodoaniline, with a perfluoroalkyl iodide (24), such as CF3(CF2)nI (e.g. N. Yoshino et. al., Bull Chem Soc Jpn, 1992;65:2141).
In Scheme 8, Step B, the desired anilines (2e) are prepared from perfluoroalkyl anilines (2d) by reductive removal of the benzylic fluorine atoms with a suitable reducting agent, such as LiAlH4 (Tetrahedron Letters, 1996;37:4655).
The anilines (5) wherein R4 is xe2x80x94(CH2)nCO2R6, n is 1, and R6 is hydrogen or ethyl are commercially available or can be prepared by one of skill in the art. The anilines (5) wherein R4 is xe2x80x94(CH2)nCO2R6, n is 1, and R6 is methyl are prepared from esterification of the commercially available corresponding aniline with diazomethane. The aniline (5) wherein R4 is xe2x80x94(CH2)nCO2R6, n is 2, and R6 is hydrogen is commercially available or can be prepared by one of skill in the art. The anilines (5) wherein R4 is xe2x80x94(CH2)nCO2R6, n is 2, and R6 is methyl or ethyl are prepared from esterification of the commercially available corresponding aniline with diazomethane. 
The anilines of Scheme 9 are used in the preparation of compounds of formula I wherein R4 is xe2x80x94(CH2)nCO2R6, n is 0, and R6 is methyl. The aniline where R2 is hydrogen is commercially available and the aniline where R2 is fluorine can be prepared according to literature procedures available to one of ordinary skill in the art.
Provided by the present invention are compounds of formula I wherein:
W is 
xe2x80x83; or
W is 
xe2x80x83; or
W is 
xe2x80x83or
W is 
xe2x80x83;
R2 is hydrogen, fluorine or chlorine; or R2 is hydrogen or fluorine; or R2 is hydrogen; or R2 is fluorine;
R3 is fluorine;
R4 is xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3, xe2x80x94(CH2)nCO2R6 or C(O)CH3 alkyl; or R4 is xe2x80x94(CH2)nCO2R6 where n is 0 and R6 is hydrogen or methyl; R4 is CO2H; or R4 is CO2CH3; or R4 is C(O)CH3; or R4 is xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3; or
R5 is hydrogen.
Also provided by the present invention are compounds of Formula 
wherein 
xe2x80x83W is
R2 is hydrogen, fluorine, or chlorine;
R3 is fluorine;
R4 is xe2x80x94CH2S(CH2)m(CH3), xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3, (Z)xe2x80x94CHCHCH2OCH3, xe2x80x94(CH2)nCO2R6, xe2x80x94(CF2)pCF3, xe2x80x94CH2(CF2)qCF3, xe2x80x94(CH2)mCF(CF3)2, xe2x80x94CH(CF3)2, xe2x80x94CF2CF(CF3)2, xe2x80x94C(CF3)3, xe2x80x94(Z)xe2x80x94CHCHxe2x80x94(CH2)qNHCH3, or (Z)xe2x80x94CHCHxe2x80x94(CH2)qN(CH3)2, or C(O)CH3;
n is 0;
R5 is hydrogen;
R6 is hydrogen or methyl;
and pharmaceutically acceptable salts thereof.
Also provided by the present invention are compounds of Formula 
wherein 
xe2x80x83W is;
R2 is hydrogen, fluorine, or chlorine;
R3 is fluorine;
R4 is xe2x80x94Cxe2x89xa1Cxe2x80x94CH2OCH3, xe2x80x94(CH2)nCO2R6 or C(O)CH3;
n is 0;
R5 is hydrogen;
R6 is hydrogen or methyl;
and pharmaceutically acceptable salts thereof.
Particularly, the present invention provides the compounds which are
Methyl 4-[[2,3-difluoro-6-[[(2-hydroxyethoxy)amino]carbonyl]-phenyl]amino]benzoate;
4-[[2,3-Difluoro-6-[[(2-hydroxyethoxy)amino]carbonyl]phenyl]-amino]benzoic acid;
3,4-Difluoro-2-[[2-fluoro-4-(3-methoxy-1-propynyl)phenyl]-amino]-N-(2-hydroxyethoxy)benzamide;
Methyl 4-(2,3-difluoro-6-{[(2-hydroxyethoxy)amino]carbonyl}anilino)-3-fluorobenzoate;
4-(2,3-Difluoro-6-{[(2-hydroxyethoxy)amino]carbonyl}anilino)-3-fluorobenzoic acid;
2-(4-Acetyl-2-chloro-phenylamino)-3,4-difluoro-N-(2-hydroxyethoxy)-benzamide and
2-{2-fluoro-4-[(methylamino)carbonyl]anilino}-3,4-difluoro-N-(2-hydroxyethoxy)benzamide.
Compounds of the present invention include, but are not limited to the following compounds:
Methyl 4-[[2,3-difluoro-6-[[(2-hydroxyethoxy)amino]carbonyl]-phenyl]amino]benzoate;
4-[[2,3-Difluoro-6-[[(2-hydroxyethoxy)amino]carbonyl]phenyl]-amino]benzoic acid; and
3,4-Difluoro-2-[[2-fluoro-4-(3-methoxy-1-propynyl)phenyl]amino]-N-(2-hydroxyethoxy)benzamide.
Also provided by the present invention are compounds which include, but are not limited to the following compounds:
3,4-Difluoro-2-(2-fluoro-4-methylsulfanylmethyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
2-(4-Ethylsulfanylmethyl-2-fluoro-phenylamino)-3,4-difluoro-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(3-methylamino-prop-1-ynyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(3-methoxy-prop-1-ynyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-((Z)-3-methoxy-propenyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluorobenzoic acid;
4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluorobenzoic acid methyl ester;
4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluorobenzoic acid ethyl ester;
{4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluoro-phenyl}-acetic acid;
{4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluoro-phenyl}-acetic acid methyl ester;
{4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluoro-phenyl}-acetic acid ethyl ester;
3-{4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-2-fluoro-phenyl}-propionic acid;
3-{4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluoro-phenyl)-propionic acid methyl ester;
3-(4-[2,3-Difluoro-6-(2-hydroxy-ethoxycarbamoyl)-phenylamino]-3-fluoro-phenyl}-propionic acid ethyl ester;
3,4-Difluoro-2-(2-fluoro-4-pentafluoroethyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-(2-fluoro-4-heptafluoropropyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-(2-fluoro-4-nonafluorobutyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-(2-fluoro-4-undecafluoropentyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-(2-fluoro-4-tridecafluorohexyl-phenylamino)-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(2,2,3,3,3-pentafluoro-propyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(2,2,3,3,4,4,4-heptafluoro-butyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(2,3,3,3-tetrafluoro-2-trifluoromethyl-propyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(1,2,2,2-tetrafluoro-1-trifluoromethyl-ethyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
2-(4-Acetyl-2-fluoro-phenylamino)-3,4-difluoro-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(2,2,2-trifluoro-1-trifluoromethyl-ethyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(1,1,2,3,3,3-hexafluoro-2-trifluoromethyl-propyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
3,4-Difluoro-2-[2-fluoro-4-(2,2,2-trifluoro-1,1-bis-trifluoromethyl-ethyl)-phenylamino]-N-(2-hydroxy-ethoxy)-benzamide;
Methyl 4-(2,3-difluoro-6-{[(2-hydroxyethoxy)amino]carbonyl}anilino)-3-fluorobenzoate;
4-(2,3-Difluoro-6-{[(2-hydroxyethoxy)amino]carbonyl}anilino)-3-fluorobenzoic acid;
2-(4-Acetyl-2-chloro-phenylamino)-3,4-difluoro-N-(2-hydroxy-ethoxy)-benzamide and
2-{2-fluoro-4-[(methylamino)carbonyl]anilino}-3,4-difluoro-N-(2-hydroxyethoxy)benzamide.
As used herein, the term xe2x80x9cpatientxe2x80x9d refers to any warm-blooded animal such as, but not limited to, a human, horse, dog, guinea pig, or mouse. Preferably, the patient is human.
The term xe2x80x9ctreatingxe2x80x9d for purposes of the present invention refers to treatment, prophylaxis or prevention, amelioration or elimination of a named condition once the condition has been established.
Selective MEK 1 or MEK 2 inhibitors are those compounds which inhibit the MEK 1 or MEK 2 enzymes, respectively, without substantially inhibiting other enzymes such as MKK3, PKC, Cdk2A, phosphorylase kinase, EGF, and PDGF receptor kinases, and C-src. In general, a selective MEK 1 or MEK 2 inhibitor has an IC50 for MEK 1 or MEK 2 that is at least one-fiftieth ({fraction (1/50)}) that of its IC50 for one of the above-named other enzymes. Preferably, a selective inhibitor has an IC50 that is at least {fraction (1/100)}, more preferably {fraction (1/500)}, and even more preferably {fraction (1/1000)}, {fraction (1/5000)}, or less than that of its IC50 or one or more of the above-named enzymes.
The disclosed compositions are useful as both prophylactic and therapeutic treatments for diseases or conditions related to the hyperactivity of MEK, as well as diseases or conditions modulated by the MEK cascade. Examples include, but are not limited to, stroke, septic shock, heart failure, osteoarthritis, rheumatoid arthritis, organ transplant rejection, and a variety of tumors such as ovarian, lung, pancreatic, brain, prostatic, and colorectal.
The invention further relates to a method for treating proliferative diseases, such as cancer, restenosis, psoriasis, autoimmune disease, and atherosclerosis. Other aspects of the invention include methods for treating MEK-related (including Ras-related) cancers, whether solid or hematopoietic. Examples of cancers include brain, breast, lung, such as non-small cell lung, ovarian, pancreatic, prostate, renal, colorectal, cervical, acute leukemia, and gastric cancer. Further aspects of the invention include methods for treating or reducing the symptoms of xenograft (cell(s), skin, limb, organ or bone marrow transplant) rejection, osteoarthritis, rheumatoid arthritis, cystic fibrosis, complications of diabetes (including diabetic retinopathy and diabetic nephropathy), hepatomegaly, cardiomegaly, stroke (such as acute focal ischemic stroke and global cerebral ischemia), heart failure, septic shock, asthma, Alzheimer""s disease, and chronic or neuropathic pain. Compounds of the invention are also useful as antiviral agents for treating viral infections such as HIV, hepatitis (B) virus (HBV), human papilloma virus (HPV), cytomegalovirus (CMV), and Epstein-Barr virus (EBV). These methods include the step of administering to a patient in need of such treatment, or suffering from such a disease or condition, a therapeutically effective amount of a disclosed compound of Formula I or pharmaceutical composition thereof.
The term xe2x80x9cchronic painxe2x80x9d for purposes of the present invention includes, but is not limited to, neuropathic pain, idiopathic pain, and pain associated with chronic alcoholism, vitamin deficiency, uremia, or hypothyroidism. Chronic pain is associated with numerous conditions including, but not limited to, inflammation, arthritis, and postoperative pain.
As used herein, the term xe2x80x9cneuropathic painxe2x80x9d is associated with numerous conditions which include, but are not limited to, inflammation, postoperative pain, phantom limb pain, burn pain, gout, trigeminal neuralgia, acute herpetic and postherpetic pain, causalgia, diabetic neuropathy, plexus avulsion, neuroma, vasculitis, viral infection, crush injury, constriction injury, tissue injury, limb amputation, arthritis pain, and nerve injury between the peripheral nervous system and the central nervous system.
The invention also features methods of combination therapy, such as a method for treating cancer, wherein the method further includes providing radiation therapy or chemotherapy, for example, with mitotic inhibitors such as a taxane or a vinca alkaloid. Examples of mitotic inhibitors include paclitaxel, docetaxel, vincristine, vinblastine, vinorelbine, and vinflunine. Other therapeutic combinations include a MEK inhibitor of the invention and an anticancer agent such as cisplatin, 5-fluorouracil or 5-fluoro-2-4(1H,3H)-pyrimidinedione (5FU), flutamide, and gemcitabine.
The chemotherapy or radiation therapy may be administered before, concurrently, or after the administration of a disclosed compound according to the needs of the patient.
Those skilled in the art will be able to determine, according to known methods, the appropriate therapeutically-effective amount or dosage of a compound of the present invention to administer to a patient, taking into account factors such as age, weight, general health, the compound administered, the route of administration, the type of pain or condition requiring treatment, and the presence of other medications. In general, an effective amount or a therapeutically-effective amount will be between about 0.1 and about 1000 mg/kg per day, preferably between about 1 and about 300 mg/kg body weight, and daily dosages will be between about 10 and about 5000 mg for an adult subject of normal weight. Commercially available capsules or other formulations (such as liquids and film-coated tablets) of 100, 200, 300, or 400 mg can be administered according to the disclosed methods.
The compounds of the present invention are preferably formulated prior to administration. Therefore, another aspect of the present invention is a pharmaceutical composition comprising a compound of Formula I and a pharmaceutically acceptable carrier. In making the compositions of the present invention, the active ingredient, such as a compound of Formula I, will usually be mixed with a carrier, or diluted by a carrier or enclosed within a carrier. Dosage unit forms or pharmaceutical compositions include tablets, capsules, pills, powders, granules, aqueous and nonaqueous oral solutions and suspensions, and parenteral solutions packaged in containers adapted for subdivision into individual doses.
Dosage unit forms can be adapted for various methods of administration, including controlled release formulations, such as subcutaneous implants. Administration methods include oral, rectal, parenteral (intravenous, intramuscular, and subcutaneous), intracisternal, intravaginal, intraperitoneal, intravesical, local (drops, powders, ointments, gels, or cream), and by inhalation (a buccal or nasal spray).
Parenteral formulations include pharmaceutically acceptable aqueous or nonaqueous solutions, dispersion, suspensions, emulsions, and sterile powders for the preparation thereof. Examples of carriers include water, ethanol, polyols (propylene glycol, polyethylene glycol), vegetable oils, and injectable organic esters such as ethyl oleate. Fluidity can be maintained by the use of a coating such as lecithin, a surfactant, or maintaining appropriate particle size. Carriers for solid dosage forms include (a) fillers or extenders, (b) binders, (c) humectants, (d) disintegrating agents, (e) solution retarders, (f) absorption acccelerators, (g) adsorbants, (h) lubricants, (i) buffering agents, and (j) propellants.
Compositions may also contain adjuvants such as preserving, wetting, emulsifying, and dispensing agents; antimicrobial agents such as parabens, chlorobutanol, phenol, and sorbic acid; isotonic agents such as a sugar or sodium chloride; absorption-prolonging agents such as aluminum monostearate and gelatin; and absorption-enhancing agents.
The following examples represent typical syntheses of the compounds of the present invention as described generally above. These examples are illustrative only and are not intended to limit the invention in any way. The reagents and starting materials are readily available to one of ordinary skill in the art. 
Step A: Preparation of 1,2:5,6-di-O-isopropylidene-D-mannitol
To a stirring suspension of D-Mannitol (1.82 g, 10.0 mmol) in tetrahydrofuran (21 mL) and dimethylformamide (9 mL) was added p-toluenesulfonic acid monohydrate (0.02 g, 0.1 mmol,) at ambient temperature, followed by 2,2-dimethoxypropane (2.8 mL, 0.023 mol). The reaction mixture was stirred for 18 hours at room temperature, then additional 2,2-dimethoxypropane (0.3 mL, 2.4 mmol) was added. The suspension was heated to 40xc2x0 C. to 45xc2x0 C., and stirred for 2 hours. Sodium bicarbonate (1.8 g, 0.016 mol) was added to neutralize the acid and the mixture was stirred for 30 minutes. The excess Na2CO3 was filtered and washed with tetrahydrofuran (5 mL). The filtrate was concentrated. To the remaining light yellow oil was added toluene (15 mL) and the mixture was stirred at 3xc2x0 C. to 5xc2x0 C. until a light-yellow gelatinous solid formed. The solid was filtered and washed with hexane (2xc3x975 mL). The product was dried in a vacuum oven for 18 hours to give 1,2:5,6-di-O-isopropylidene-D-mannitol (1.24 g, 47.3%) as an off-white solid, mp 110-113xc2x0 C. 
Step B: Preparation of (S)-(+)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol
To a solution of the product of Preparation 1, Step A, 1,2:5,6-di-O-isopropylidene-D-mannitol (50 g, 0.191 mol), in water (700 mL), was added solid sodium bicarbonate (20 g). The resultant solution was stirred until all the solid dissolved, and then cooled in an ice-water bath. Solid sodium periodate (81.5 g, 0.381 mol) was slowly added to the solution portionwise. Gas evolution observed. The white mixture was stirred at ambient temperature for 2 hours. Solid sodium chloride (30 g) was added, and the mixture was stirred for 15 minutes. The white solid was filtered. The filtrate was cooled in an ice-water bath. Solid sodium borohydride was added slowly. Gas bubble evolved. The mixture was warmed to ambient temperature, and stirred overnight. The milky mixture turned to a clear solution. The aqueous solution was extracted with dichloromethane (3xc3x97). The organic solution was washed with brine, and dried over magnesium sulfate. The solvent was removed in vacuo to give (S)-(+)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol as a colorless oil, which was dried under high vacuum at ambient temperature overnight, 34.82 g (60%); MS (APCI+)=133 (M++1).
Step C: Preparation of (R)-2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione
A 3-L round-bottomed flask equipped with mechanical stirrer and additional funnel was charged with N-hydroxyphthalimide (68.0 g, 0.416 mol) and tetrahydrofuran (1.2 L) under nitrogen atmosphere. To this solution was added triphenylphosphine (109.2 g, 0.416 mol) and the product of Preparation 1, Step B, (S)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol (55.0 g, 0.416 mol). The mixture was cooled to 3xc2x0 C. to 5xc2x0 C. and diethyl azodicarboxylate (85.2 mL, 0.541 mol) was added dropwise, while keeping the inner temperature below 15xc2x0 C. The reaction mixture was warmed to ambient temperature, and stirred for 18 hours. The tetrahydrofuran was evaporated under reduced pressure. To the remaining orange solid was added dichloromethane (0.5 L) and the mixture was stirred for 1 hour. The white solid (Ph3PO) was filtered and washed with dichloromethane (0.1 L). The solvent was removed and ethanol (0.5 L) was added to the resulting solid. The mixture was stirred for 2 hours at 3xc2x0 C. to 5xc2x0 C. The white solid was filtered, washed with a small amount of cold ETOH, and dried in vacuum oven at 40xc2x0 C. to give (R)-2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione (112.5 g, 97%) as a white solid: 1H NMR (CDCl3): xcex4 1.33 (s, 3H), 1.99 (s, 3H), 3.96 (m, 1H), 4.15 (m, 2H), 4.30 (m, 1H), 4.48 (m, 1H), 7.59 (m, 2H), 7.84 (m, 2H); MS (APCI+)=278 (M++1). 
Step D: Preparation of (R)-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-hydroxylamine
To a stirring solution of the product of Preparation 1, Step C, (R)-2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione (74.9 g, 0.27 mol) in dichloromethane (480 mL) at 3xc2x0 C. to 5xc2x0 C. was added methylhydrazine (15.8 mL, 0.29 mol) dropwise. The color of the suspension turned from yellow to white. The cooling bath was removed and the mixture was stirred for 2 hours at ambient temperature. The resulting suspension was concentrated on a rotary evaporator. To the white solid was added ether (0.5 L) and the resulting mixture was stirred for 1.5 hours at ambient temperature. The white precipitate was filtered and washed with ether (0.2 L). The filtrate was concentrated on rotary evaporator to give (R)-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-hydroxylamine (39.0 g, 98.3%): 1H NMR (CDCl3): xcex4 1.35 (s, 3H), 1.42 (s, 3H), 3.73 (m, 3H), 4.05 (m, 1H), 4.33 (m, 1H), 5.39 (m, 2H); MS (APCI+)=148.1 (M++1). 
Step A: Preparation of L-gulonic xcex3-lactone
To a solution of L-ascorbic acid (83.9 g, 0.477 mol) in water (600 mL) was added Pd/C (10%, 8.3 g). The mixture was subjected to hydrogenation in a Parr hydrogenator at 48 psi, 18xc2x0 C. for 62 hours. The reaction mixture was filtered and the filtrate was concentrated in vacuo to afford L-gulonic xcex3-lactone (81.0 g, 96%) as an off-white solid, after drying at 50xc2x0 C. in a vacuum oven for 18 hours: mp 182-184xc2x0 C.
Step B: Preparation of 5,6-isopropylidene-L-gulonic Acid xcex3-lactone
The product of Preparation 2, Step A, L-gulonic xcex3-lactone (25.0 g, 140.3 mmol) was dissolved in mixture of tetrahydrofuran (140 mL) and dimethylformamide (200 mL). p-Toluenesulfonic acid monohydrate (2.67 g, 14.0 mmol) was added and the reaction mixture was cooled to 0xc2x0 C. to 5xc2x0 C. in an ice-water bath. 2,2-Dimethoxypropane (22.4 mL, 182.4 mmol) was added dropwise and the reaction mixture was stirred at ambient temperature for 18 hours. The mixture was neutralized with solid sodium carbonate (24.0 g) and stirred for 1 hour. The solid was filtered and washed with tetrahydrofuran. The THF was removed under vacuo, and DMF by distillation under high vacuum. The resulting orange solid was triturated with toluene (300 mL), filtered, washed with toluene (20 mL), and dried in a vacuum oven at 40xc2x0 C. for 3 days, to yield 5,6-isopropylidene-L-gulonic acid xcex3-lactone (28.9 g, 94%) as a pale orange solid: mp 155-158xc2x0 C.; MS (APCI+)=219.0 (M++1). 
Step C: Preparation of (R)-(+)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol
To a stirring suspension of the product of Preparation 2, Step B, 5,6-O-isopropylidene-L-gulono-1,4-lactone (15.16 g, 69.5 mmol) in water (0.3 L) was added solid sodium periodate in small portions at 3xc2x0 C. to 5xc2x0 C. The pH of the mixture was adjusted to 5.5 with 1N aqueous sodium hydroxide. The suspension was stirred for 2 hours at ambient temperature, then saturated with sodium chloride (20.0 g) and filtered. To the filtrate, at 3xc2x0 C. to 5xc2x0 C., was added sodium borohydride (10.5 g, 0.278 mol) in small portions. The reaction mixture was stirred for 18 hours at ambient temperature. Acetone (100 mL) was added to destroy the excess of sodium borohydride, and the stirring was continued for 30 minutes. The acetone was removed under reduced pressure, and the aqueous residue was extracted with dichloromethane (3xc3x97300 mL) and EtOAc (3xc3x97300 mL). The combined organic layers were dried over magnesium sulfate, filtered, and evaporated to give (R)-(+)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol (5.07 g, 55.7%) as a colorless clear liquid: MS (APCI+)=132.9 (M++1).
Step D: Preparation of (S)-2-(2,2-Dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione
A 3-L round-bottomed flask equipped with mechanical stirrer and additional funnel was charged with N-hydroxyphthalimide (68.0 g, 0.416 mol) and tetrahydrofuran (1.2 L) under nitrogen atmosphere. To this solution was added triphenylphosphine (109.2 g, 0.416 mol) and (R)-(2,2-dimethyl-[1,3]dioxolan-4-yl)-methanol (55.0 g, 0.416 mol). The mixture was cooled to 3xc2x0 C. to 5xc2x0 C. and diethyl azodicarboxylate (85.2 mL, 0.541 mol) was added dropwise, while keeping the inner temperature below 15xc2x0 C. The reaction mixture was warmed to ambient temperature, and stirred for 18 hours. The tetrahydrofuran was evaporated under reduced pressure. To the remaining orange solid was added dichloromethane (0.5 L) and the mixture was stirred for 1 hour. The white solid (Ph3PO) was filtered and washed with dichloromethane (0.1 L). The solvent was removed and ethanol (0.5 L) was added to the resulting solid. The mixture was stirred for 2 hours at 3xc2x0 C. to 5xc2x0 C. The white solid was filtered, washed with a small amount of cold EtOH and dried in vacuum oven at 40xc2x0 C. to give (S)-2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione (112.5 g, 97%) as a white solid: 1H NMR (CDCl3): xcex4 1.33 (s, 3H), 1.99 (s, 3H), 3.96 (m, 1H), 4.15 (m, 2H), 4.30 (m, 1H), 4.48 (m, 1H), 7.59 (m, 2H), 7.84 (m, 2H); MS (APCI+)=278 (M++1).
Step E: Preparation of (S)-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-hydroxylamine
To a stirring solution of (S)-2-(2,2-dimethyl-[1,3]dioxolan-4-ylmethoxy)-isoindole-1,3-dione (74.9 g, 0.27 mol) in dichloromethane (480 mL) at 3xc2x0 C. to 5xc2x0 C. was added methylhydrazine (15.8 mL, 0.29 mol) dropwise. The color of the suspension turned from yellow to white. The cooling bath was removed and the mixture was stirred for 2 hours at ambient temperature. The resulting suspension was concentrated on a rotary evaporator. To the white solid was added ether (0.5 L) and the resulting mixture was stirred for 1.5 hours at ambient temperature. The white precipitate was filtered and washed with ether (0.2 L). The filtrate was concentrated on rotary evaporator to give (S)-(2,2-dimethyl-[1,3]dioxolan-4-ylmethyl)-hydroxylamine (39.0 g, 98.3%): 1H NMR (CDCl3): xcex4 1.35 (s, 3H), 1.42 (s, 3H), 3.73 (m, 3H), 4.05 (m, 1H), 4.33 (m, 1H), 5.39 (m, 2H); MS (APCI+)=148.1 (M++1). 
Step A: Preparation of 2-(2,2-dimethyl-[1,3]dioxan-5-yloxy)-isoindole-1,3-dione 2,2-Dimethyl-[1,3]dioxan-5-ol was Prepared as Described Previously
(Forbes, D. C. et. al. Synthesis, 1998;6:879-82). 1H NMR (400 MHz; DMSO-d6) xcex4 4.91 (d, 1H, J=5.1), 3.70-3.75 (m, 2H), 3.41-3.46 (m, 3H), 1.30 (s, 3H), 1.24 (s, 3H); MS (APCI+)=132.9. To a stirring solution of 2,2-dimethyl-[1,3]dioxan-5-ol (1.50 g, 11.35 mmol), N-hydroxyphthalimide (1.85 g, 11.35 mmol) and triphenylphosphine (2.98 g, 11.35 mmol) in anhydrous tetrahydrofuran (30 mL) at 0xc2x0 C. was added diethyl azodicarboxylate (2.3 mL, 14.75 mmol). The resultant solution was allowed to warm to room temperature. After stirring for 3 hours, the mixture was concentrated in vacuo and charged with chloroform affording white solids. The solids were filtered off and filtrate was collected and concentrated. The residue was purified via silica column chromatography (4:1 hexanes/ethyl acetate) affording 2-(2,2-dimethyl-[1,3]dioxan-5-yloxy)-isoindole-1,3-dione as clear crystals (1.74 g, 55% over 2 steps): 1H NMR (400 MHz; DMSO-d6) xcex4 7.83 (s, 4H), 4.11-4.12 (m, 1H), 4.04-4.09 (m, 2H), 3.92-3.96 (m, 2H), 1.32 (s, 3H), 1.25 (s, 3H); MS (APCI+)=278.0.
Step B: Preparation of O-(2,2-dimethyl-[1,3]dioxan-5-yl)-hydroxylamine
To a stirring solution of the product of Example 13, Step A, 2-(2,2-dimethyl-[1,3]dioxan-5-yloxy)-isoindole-1,3-dione (1.72 g, 6.20 mmol) in dichloromethane (15 mL) at 0xc2x0 C. under nitrogen was added methylhydrazine (0.36 mL, 6.82 mmol) and allowed to warm to room temperature. After stirring for 2 hours the reaction mixture was concentrated in vacuo and charged with diethylether. The solids were filtered off and the filtrate was collected and concentrated to afford O-(2,2-dimethyl-[1,3]dioxan-5-yl)-hydroxylamine as a yellow oil (0.97 g, 100%). 1H NMR (400 MHz; DMSO-d6) xcex4 5.98 (bs, 2H), 3.84-3.87 (m, 2H), 3.66-3.68 (m, 2H), 3.30-3.35 (m, 1H), 1.29 (s, 3H), 1.22 (s, 3H); MS (APCI+)=147.9.